Project Summary/ Abstract Defining the genetic and epigenetic abnormalities that drive cellular transformation has provided insights into potential therapeutic approaches to cancer. However, the myriad of genetic changes that accompany specific types of cancer makes it difficult to find therapeutic approaches applicable to the bulk of patients with a given disease. We have identified an oncogenic role for the epigenetic enzyme ?co-activator associated arginine methyltransferase 1? (CARM1) in AML, and a dependency of AML cells, but not normal hematopoietic stem cells, on CARM1 activity. CARM1 methylates a variety of histone and non-histone substrates and targeting its activity, via small molecule inhibitors, knockdown (KD) or knockout (KO) approaches, triggers AML cell differentiation and death. The proposed studies represent a conceptual shift from targeting specific mutations found in AML to developing therapies that target chromatin modifiers and transcriptional regulators that are essential to large numbers of AML patients. To advance this new paradigm we have proposed the following specific aims: Aim 1: We will define the molecular basis for the differentiation-promoting effects of CARM1 inhibition on AML cells, identifying the critical CARM1-interacting proteins, substrates and target genes that regulate myeloid differentiation, proliferation and survival. We will use state-of- the-art techniques including the BioID2 system, mass-spectrometry, RNA-Seq and ChIP-Seq. Aim 2: We will determine the mechanisms that control the level of CARM1 activity in AML cells, defining how changes in CARM1 mRNA levels, protein levels, and post- transcriptional modifications, including phosphorylation, affect CARM1 activity in AML cells. Aim 3: Determine the basis for the unique sensitivity of AML cells to CARM1 inhibition or KD. We will also define CARM1 inhibitor resistance mechanisms, and test CARM1 inhibitor-containing combination therapies, using human AML cell lines and primary AML samples and NOD/SCID mice engrafted with human AML cells. These studies will help establish the importance of CARM1 in the pathogenesis of AML, and advance our understanding of how CARM1 inhibitors can be used as part of an effective and novel ?epigenetic-targeted? strategy for treating cancer.